GSPE Treatment Research Articles

Enhanced osteogenesis on proantocyanidin-loaded date palm endocarp cellulosic matrices: A novel sustainable approach for guided bone regeneration

Developing inexpensive, biocompatible natural scaffolds that can support the differentiation and proliferation of stem cells has been recently emphasized by the research community to faster obtain the FDA approvals for regenerative medicine. In this regard, plant-derived cellulose materials are a novel class of sustainable scaffolding materials with high potentials for bone tissue engineering (BTE). However, low bioactivity of the plant-derived cellulose scaffolds restricts cell proliferation and cell differentiation. This limitation can be addressed though surface-functionalization of cellulose scaffolds with natural antioxidant polyphenols, e.g., grape seed proanthocyanidin (PCA)-rich extract (GSPE). Despite the various merits of GSPE as a natural antioxidant, its impact on the proliferation and adhesion of osteoblast precursor cells, and on their osteogenic differentiation is an as-yet unknown issue. Here, we investigated the effects of GSPE surface functionalization on the physicochemical properties of decellularized date (Phoenix dactyliferous) fruit inner layer (endocarp) (DE) scaffold. In this regard, various physiochemical characteristics of the DE-GSPE scaffold such as hydrophilicity, surface roughness, mechanical stiffness, porosity, and swelling, and biodegradation behavior were compared with those of the DE scaffold. Additionally, the impact of the GSPE treatment of the DE scaffold on the osteogenic response of human mesenchymal stem cells (hMSCs) was thoroughly studied. For this purpose, cellular activities including cell adhesion, calcium deposition and mineralization, alkaline phosphatase (ALP) activity, and expression levels of bone-related genes were monitored. Taken together, the GSPE treatment enhanced the physicochemical and biological properties of the DE-GSPE scaffold, thereby raising its potentials as a promising candidate for guided bone regeneration.

Grape seed proanthocyanidin extract targets p66Shc to regulate mitochondrial biogenesis and dynamics in diabetic kidney disease.

In vivo, rats were intraperitoneally injected with streptozotocin (STZ) and treated with GSPE. Biochemical changes, mitochondrial morphology, the ultrastructure of nephrons, and protein expression of mitochondrial biogenesis (SIRT1, PGC-1α, NRF1, TFAM) and dynamics (DRP1, MFN1) were determined. In vitro, HK-2 cells were transfected with p66Shc and treated with GSPE to evaluate changes in cell apoptosis, reactive oxygen species (ROS), mitochondrial quality, the protein expression. In vivo, GSPE significantly improved the renal function of rats, with less proteinuria and a lower apoptosis rate in the injured renal tissue. Besides, GSPE treatment increased SIRT1, PGC-1α, NRF1, TFAM, and MFN1 expression, decreased p66Shc and DRP1 expression. In vitro, overexpression of p66Shc decreased the resistance of HK-2 cells to high glucose toxicity, as shown by increased apoptosis and ROS production, decreased mitochondrial quality and mitochondrial biogenesis, and disturbed mitochondrial dynamic homeostasis, ultimately leading to mitochondrial dysfunction. While GSPE treatment reduced p66Shc expression and reversed these changes. GSPE can maintain the balance between mitochondrial biogenesis and dynamics by negatively regulating p66Shc expression.

Evaluation of the Cardiac Protection Conferred by Proanthocyanidins in Grape Seeds against Development of Ehrlich Solid Tumors in Mice.

Examination of the antineoplastic effects of a range of chemical compounds is often undertaken via the transplantable tumor model of Ehrlich solid tumor (EST), which is a simulation of breast cancer. The purpose of this study was to explore how cardiac toxicity, damage, oxidative stress, and changes in the expressions of TNFα and apoptotic P53 triggered by EST could be countered with grape seed proanthocyanidins (GSPE). To that end, 50 female mice were used, with arbitrary and equal distribution into five groups, namely, the control group (G1), GSPE group (G2), EST group (G3), GSPE + EST (G4; cotreatment consisted of mice that received GSPE treatment at the beginning of EST induction over a period of 14 days), and EST + GSPE (G5; posttreatment consisted of mice with EST that received GSPE treatment for 14 days following the 14 days since the induction of EST). By comparison with the control group, the EST group had significantly higher levels of serum lactate dehydrogenase (LDH), creatine phosphokinase (CPK), creatine kinase MB (CK-MB), myoglobin, cardiac TBARS, nitric oxide (NO), total thiol and hydrogen peroxide, cardiac damage, and expression of P53 and TNFα. On the other hand, the EST group had significantly lower levels of cardiac catalase and total antioxidant (TAC) than the control group. Furthermore, better improvement in cardiac toxicity, oxidative stress, damage, apoptosis, and TNFα expressions was displayed by the cotreated (GSPE + EST) group than by the posttreated (EST + GSPE) group. This led to the conclusion that GSPE conferred cardiac protective and antioxidant effects against EST. This finding calls for more investigation on the benefits of grape seeds as adjuvant agents to prevent and treat cardiac toxicity.

Protective Effect of Proanthocyanidins in a Rat Model of Mild Intestinal Inflammation and Impaired Intestinal Permeability Induced by LPS.

Intestinal dysfunction consists of a defective barrier function, which allows the influx of luminal endotoxins, thus causing intestinal inflammation. Proanthocyanidins are natural bioactive compounds that could modulate intestinal dysfunction. This study analyzes the protective effects of proanthocyanidins in a rat model of intestinal dysfunction. To investigate the preventive effects of both high dietary (75 mgkg-1 body weight) and pharmacological (375 mgkg-1 body weight) oral doses of proanthocyanidins (GSPE), rat intestinal dysfunction is induced with LPS (i.p.). In vivo intestinal permeability (ovalbumin [OVA] assay) and systemic inflammation and endotoxemia (TNF-α and LPS plasma levels) are assessed. Intestinal inflammation and oxidative stress are determined using myeloperoxidase (MPO), cyclooxygenase-2 (COX-2) activities, and reactive oxygen species (ROS) levels, respectively. Ileal gene expression of permeability/inflammatory genes is analyzed. LPS administration induces intestinal permeability, inflammation, and oxidative stress. GSPE normalizes in vivo OVA levels. In the small intestine, the GSPE treatment decreases MPO and COX-2 activities; modulates the ileum inflammatory and permeability proteins gene expression; and in the large intestine, prevents increase of ROS levels. Proanthocyanidins, at nutritional and pharmacological doses, prevents endotoxin-induced-intestinal inflammation, permeability, and oxidative stress in rats differentially in each intestinal section. Proanthocyanidins are nutritional-therapeutic novel candidates for preventing intestinal dysfunction.

Grape Seed Proanthocyanidin Extract Alleviates AflatoxinB₁-Induced Immunotoxicity and Oxidative Stress via Modulation of NF-κB and Nrf2 Signaling Pathways in Broilers.

Aflatoxin B1 (AFB1) is a widely spread mycotoxin contaminates food and feed, causing severe oxidative stress damages and immunotoxicity. Grape seed proanthocyanidin (GSPE), a natural antioxidant with wide range of pharmacological and medicinal properties. The goal of the present study was to investigate the protective effects of GSPE against AFB1-induced immunotoxicity and oxidative stress via NF-κB and Nrf2 signaling pathways in broiler chickens. For the experiment, 240 one-day old Cobb chicks were allocated into four dietary treatment groups of six replicates (10 birds per replicate): 1. Basal diet (control); 2. Basal diet + AFB1 1mg/kg contaminated corn (AFB1); 3. Basal diet + GSPE 250 mg/kg (GSPE); 4. Basal diet + AFB1 1 mg/kg + GSPE 250 mg/kg (AFB1 + GSPE). The results showed that GSPE significantly decreased serum inflammatory cytokines TNF-α, IFN-γ, IL-1β, IL-10, and IL-6 induced by AFB1. Similarly, GSPE + AFB1 treated group revealed a significant decrease in mRNA expressions of pro-inflammatory cytokines (TNF-α, IFN-γ, IL-1β, and IL-6) in the splenic tissue compared to the AFB1 treatment group. In addition, western blotting results manifested that GSPE treatment normalized the phosphorylation of nuclear factor kappa B (p65) and the degradation of IκBα protein induced by AFB1. Furthermore, GSPE enhanced the antioxidant defense system through activating the nuclear factor-erythroid-2-related factor (Nrf2) signaling pathway. The mRNA and protein expression level of Nrf2 and its down streaming associated genes were noted up-regulated by the addition of GSPE, and down-regulated in the AFB1 group. Taken together, GSPE alleviates AFB1-induced immunotoxicity and oxidative damage by inhibiting the NF-κB and activating the Nrf2 signaling pathways in broiler chickens. Conclusively, our results suggest that GSPE could be considered as a potential natural agent for the prevention of AFB1-induced immunotoxicity and oxidative damage.

Chemo-Preventive Functions of Grape Seed Proanthocyanidin Extract against UV-Induced Intracellular Oxidative Stress and Tyrosinase Activity

Clinical studies have identified that an excessive exposure of UV light can cause oxidative stress (OS) and tyrosinase enzyme over-expression, which are associated with multiple diseases including atherosclerosis, cancers, diabetics, rheumatoid arthritis. In this study, we investigated the impact of grape seed proanthocyanidin (GSPE) on regulating OS and tyrosinase activity in human epidermal melanocytes. This study revealed that GSPE did not affect cell viability and protected cells from UV induced damage in a dose-dependent manner. 5-(-6)-Carboxy-2,7-di-chlorodihydro-fluorescein diacetate staining (i.e., a fluorescence staining for intracellular (OS)) indicated that GSPE reduced OS level caused by UV exposure. A similar trend was also confirmed by flow cytometry analysis, where GSPE down-regulated OS level. Tyrosinase analysis showed that GSPE treatment decreased tyrosinase activity. Taken all data together, GSPE may restore the cellular damage caused by excessive UV-exposure and promote skin health by reducing tyrosine generation. Clinically, GSPE could be potentially utilized for improving skin health against excessive UV exposure.

Proanthocyanidin and fish oil potent activity against cisplatin-induced renal cell cycle arrest and apoptosis in rats

Cisplatin is an effective chemotherapeutic agent that displays dose-limiting nephrotoxicity. In the present study, the efficacy of grape seed proanthocyanidin extract (GSPE: 100 mg/kg/day) and fish oil (FO: 5 mL/kg/day) against cisplatin-induced nephrotoxicity was evaluated in terms of DNA damage, histopathological changes and expression levels of molecular markers of apoptosis. The administration of cisplatin (CP) (7 mg/kg) results in an increasing percentage of S-phase, G2/M and apoptosis. Furthermore, CP induces apoptosis as indicated by an elevation of renal caspase-3 and reduction in the expression of BCL-2. In addition to occurred renal histopathological changes as manifested by tubular degeneration, degenerative glomerulus, necrotic tubular cells, and cell debris. On the other hand, the administration of GSPE or FO pre-cisplatin treatment can be ameliorated the current DNA cell cycle alterations by the restoration of expression of proteins related to apoptosis and reduced the undesirable renal histopathological changes. So, it can be concluded that the consumption of GSPE or FO might be useful for minimizing nephrotoxicity caused by cisplatin chemotherapy through their anti-apoptotic and antioxidant properties.

Effect of grape seed proanthocyanidin on mitochondrial injury in hippocampus and cognition impairment in a rat model of obstructive sleep apnea hypoxia

Objective To investigate the effects of grape seed proanthocyanidin(GSPE) on mitochondrial injury in hippocampus and learning-memory impairment after obstructive sleep apnea hypoxia in rats. Methods Male SD rats(n=80) were randomly divided into control group, model group, low dose of GSPE treatment group and high dose of GSPE treatment group.Rats in control group were exposed in air, the model group were suffered from intermittent hypoxia conditions(50 ml/L, 8-hour-intermittent hypoxia everyday, and the duration of experiment 2 and 6 weeks, respectively). Mitochondrion pathology in hippocampal region was observed using electron microscope; malondialdehyde (MDA) contents and superoxide dismutase activity were detected by colorimetry and apoptotic cells was measured by TUNEL method. The cognitive function of rats in each group was assessed with the Morris water maze (MWM). Results After hypoxia, mitochondrion was significantly injured.The MDA contents were increased(79.86±2.52, 88.26±2.86) and SOD level decreased(70.67±6.70, 64.26±7.86). The number of neural apoptotic cells was significantly enhanced (9.68±0.79, 15.9±2.92). MWM test showed that the escaping latency was prolonged and the frequency of crossing the platform was decreased(P<0.05). Compared with that in the model group, low dose of GSPE decreased MDA contents (76.38±1.96, 82.16±2.02), increased SOD level(76.20±6.86, 70.58±6.86), and decreased apoptotic cells(6.60±0.69, 9.54±1.36). MWM test showed that the escaping latency was shortened and the frequency of crossing the platform was increased in GSPE treatment groups(P<0.05). Compared with low dose of GSPE, high dose of GSPE decreased MDA contents increased SOD level and decreased apoptotic cells.MWM test showed that the escaping latency was shortened and the frequency of crossing the platform was increased (P<0.05). Conclusion GSPE can attenuate mitochondrial injury and improve learning-memory function after obstructive sleep apnea hypoxia. Key words: Sleep apnea; Learning memory; Mitochondrion; Grape seed proanthocyanidin

Effects of grape seed procyanidin extract over low-grade chronic inflammation of obese Zucker fa/fa rats

Obesity is associated with chronic inflammation and is related to the accumulation and release of several cytokines from adipose tissue. It has been reported that a grape seed extract (GSPE) containing large amounts of procyanidins, phenolic compounds widely distributed in food, has anti-inflammatory effects in vivo against chronic inflammation related to obesity induced by the diet. In this study, several Zucker fa/fa rats were used as the obese control group (n=10) and received vehicle treatments, whereas other Zucker fa/fa rats (n=10) were treated with a moderate dose of 35mg/kg∗day of GSPE for a period of 10weeks. Importantly, GSPE treatment significantly decreased the average level of C-reactive protein (CRP) at 5weeks of treatment. However, at the end of the experiment (10weeks of treatment), GSPE could not decrease any of the pro-inflammatory markers tested such as MCP-1, TNF-α or CRP. On the contrary, GSPE showed anti-inflammatory effects on mesenteric adipose tissue, as GSPE treatment down-regulated the expression of iNos and Il-6 and up-regulated the expression of Adiponectin in the adipocytes. In conclusion, a moderate dose of GSPE exerts an anti-inflammatory effect in genetically obese rats, modulating the expression of several genes in the adipose tissue and being the adipocyte a target cell of procyanidins. However, this dose could not counteract the pro-inflammatory state at systemic level.

Proanthocyanidins from grape seeds modulates the nuclear factor-kappa B signal transduction pathways in rats with TNBS-induced recurrent ulcerative colitis

The aim of this study was to elucidate the molecular mechanisms involved in the therapeutic effects of proanthocyanidins from grape seeds (GSPE) on recurrent ulcerative colitis (UC) in rats. GSPE in doses of 100, 200, and 400mg/kg were intragastrically administered per day for 7days after recurrent colitis was twice-induced by TNBS. The levels of GSH, as well as the activity of GSH-Px and SOD in colon tissues were measured by biochemical methods. The expression levels of tumor necrosis factor-α (TNF-α) and the nuclear translocation levels of nuclear factor-kappa B (NF-κB) in the colon tissues were measured by enzyme-linked immunosorbent assay methods. Western blotting analysis was used to determine the protein expression levels of inhibitory kappa B-alpha (IκBα), inhibitor kappa B kinase (IKKα/β), phosphorylated IκBα and phosphorylated IKKα/β. GSPE treatment was associated with a remarkable increased the activity of GSH-Px and SOD with GSH levels in TNBS-induced recurrent colitis rats as compared to the model group. GSPE also significantly reduced the expression levels of TNF-α, p-IKKα/β, p-IκBα and the translocation of NF-κB in the colon mucosa. GSPE exerted a protective effect on recurrent colitis in rats by modifying the inflammatory response and promoting damaged tissue repair to improve colonic oxidative stress. Moreover, GSPE inhibited the TNBS-induced inflammatory of recurrent colitis though blocking NF-κB signaling pathways.

Effects of proanthocyanidins from grape seed on treatment of recurrent ulcerative colitis in rats

The aim of the present study was to investigate the therapeutic effect and mechanism of proanthocyanidins from grape seed (GSPE) in the treatment of recurrent ulcerative colitis (UC) in rats. To induce recurrent colitis, rats were instilled with 2,4,6-trinitrobenzenesulfonic acid (TNBS) (80mg/kg) into the colon through the cannula in the first induced phase, and then the rats were instilled a second time with TNBS (30mg/kg) into the colon on the sixteenth day after the first induction UC. Rats were intragastrically administered GSPE (200mg/kg) per day for 7days after twice-induced colitis by TNBS. Sulfasalazine at 500mg/kg was used as a positive control drug. Rats were killed 7days after GSPE treatment. The colonic injury and inflammation were assessed by macroscopic and macroscopic damage scores, colon weight/length ratio (mg/cm), and myeloperoxidase activity. Then, superoxide dismutase, glutathione peroxidase, inducible nitric oxide synthase (iNOS) activities, and the levels of malonyldialdehyde, glutathione, and nitric oxide in serum and colonic tissues were measured. Compared with the recurrent UC group, GSPE treatment facilitated recovery of pathologic changes in the colon after induction of recurrent colitis, as demonstrated by reduced colonic weight/length ratio and macroscopic and microscopic damage scores. The myeloperoxidase and iNOS activities with malonyldialdehyde and nitric oxide levels in serum and colon tissues of colitis rats were significantly decreased in the GSPE group compared with those in the recurrent UC group. In addition, GSPE treatment was associated with notably increased superoxide dismutase, glutathione peroxidase activities, and glutathione levels of colon tissues and serum of rats. GSPE exerted a protective effect on recurrent colitis in rats by modifying the inflammatory response, inhibiting inflammatory cell infiltration and antioxidation damage, promoting damaged tissue repair to improve colonic oxidative stress, and inhibiting colonic iNOS activity to reduce the production of nitric oxide.

Therapeutic effect and mechanism of proanthocyanidins from grape seeds in rats with TNBS-induced ulcerative colitis

The aim of the study was to investigate the therapeutic effect and mechanism of proanthocyanidins from grape seeds (GSPE) in the treatment of ulcerative colitis (UC). Rats were intragastrically administered different doses of GSPE (100, 200, and 400 mg/kg) per day for 7 days after UC was twice-induced by intracolonic injection of 2,4,6-trinitrobenzene sulfonic acid (TNBS)dissolved in 50% ethanol. Sulfasalazine (SASP) at 200 mg/kg was used as a positive control drug. Macroscopic and microscopic damage scores and changes in weight/length ratio (mg/mm) of colon segments were analyzed. The levels of malonyldialdehyde (MDA), interleukin (IL)-1beta, IL-2, IL-4, and myeloperoxidase (MPO) activity in the colon tissues and MPO activity in the serum were all measured by biochemical methods or double antibody sandwich ELISA methods. Compared with the TNBS control group, GSPE treatment facilitated recovery of pathologic changes in the colon after insult with TNBS, as demonstrated by increased body weight (p < 0.01) and decreased colonic weight/length ratio (p < 0.01); GSPE also notably reduced the colonic macroscopic and microscopic damage scores (p < 0.01). The MPO activity in colon tissues and serum of rats treated with GSPE was significantly lower than that in the TNBS control group. The MDA and IL-1beta levels of colon tissues were also decreased in GSPE groups. The intestinal antiinflammatory effect of GSPE was accompanied by a significant improvement of IL-2 and IL-4 levels in the colon tissues of rats in the high-dose GSPE group (p < 0.05). Compared with the SASP group, GSPE groups had no significant difference in the therapeutic effect (p > 0.05). GSPE exerts a beneficial antiinflammatory effect in the acute phase of TNBS-induced colitis in rats by downregulating some of the mediators involved in the intestinal inflammatory response, inhibiting inflammatory cell infiltration and antioxidation damage, promoting damaged tissue repair to improve colonic oxidative stress, decreasing production of proinflammatory cytokines IL-1beta, and increasing production of antiinflammatory cytokines IL-2 and IL-4.

Dermal wound healing properties of redox-active grape seed proanthocyanidins.

Angiogenesis plays a central role in wound healing. Among many known growth factors, vascular endothelial growth factor (VEGF) is believed to be the most prevalent, efficacious, and long-term signal that is known to stimulate angiogenesis in wounds. The wound site is rich in oxidants, such as hydrogen peroxide, mostly contributed by neutrophils and macrophages. We proposed that oxidants in the wound microenvironment support the repair process. Proanthocyanidins or condensed tannins are a group of biologically active polyphenolic bioflavonoids that are synthesized by many plants. Previously we have reported that a grape seed proanthycyanidin extract containing 5000 ppm resveratrol (GSPE) potently upregulates oxidant and tumor necrosis factor-alpha inducible VEGF expression in human keratinocytes (Free Radic. Biol. Med. 31:38-42, 2001). Our current objective was to follow up on that finding and test whether GSPE influences dermal wound healing in vivo. First, using a VEGF promoter-driven luciferase reporter construct we observed that the potentiating effect of GSPE on inducible VEGF expression is at the transcriptional level. The reporter assay showed that GSPE alone is able to drive VEGF transcription. Next, two dermal excisional wounds were inflicted on the back of mice and the wounds were left to heal by secondary intention. Topical application of GSPE accelerated wound contraction and closure. GSPE treatment was associated with a more well-defined hyperproliferative epithelial region, higher cell density, enhanced deposition of connective tissue, and improved histological architecture. GSPE treatment also increased VEGF and tenascin expression in the wound edge tissue. Tissue glutathione oxidation and 4-hydroxynonenal immunostaining results supported that GSPE application enhanced the oxidizing environment at the wound site. Oxidants are known to promote both VEGF as well as tenascin expression. In summary, our current study provides firm evidence to support that topical application of GSPE represents a feasible and productive approach to support dermal wound healing.

Cellular protection with proanthocyanidins derived from grape seeds.

Grape seed proanthocyanidins have been reported to possess a broad spectrum of pharmacological and medicinal properties against oxidative stress. We have demonstrated that IH636 proanthocyanidin extract (GSPE) provides excellent protection against free radicals in both in vitro and in vivo models. GSPE had significantly better free radical scavenging ability than vitamins C, E and beta-carotene and demonstrated significant cytotoxicity towards human breast, lung and gastric adenocarcinoma cells, while enhancing the growth and viability of normal cells. GSPE protected against tobacco-induced apoptotic cell death in human oral keratinocytes and provided protection against cancer chemotherapeutic drug-induced cytotoxicity in human liver cells by modulating cell cycle/apoptosis regulatory genes such as bcl2, p53 and c-myc. Recently, the bioavailability and mechanistic pathways of cytoprotection by GSPE were examined on acetaminophen-induced hepatotoxicity and nephrotoxicity, amiodarone-induced pulmonary toxicity, doxorubicin-induced cardiotoxicity, DMN-induced immunotoxicity and MOCAP-induced neurotoxicity in mice. Serum chemistry changes, integrity of genomic DNA and histopathology were assessed. GSPE pre-exposure provided near complete protection in terms of serum chemistry changes and DNA damage, as well as abolished apoptotic and necrotic cell death in all tissues. Histopathological examination reconfirmed these findings. GSPE demonstrated concentration-/dose-dependent inhibitory effects on the drug metabolizing enzyme cytochrome P450 2E1, and this may be a major pathway for the anti-toxic potential exerted by GSPE. Furthermore, GSPE treatment significantly decreased TNFalpha-induced adherence of T-cells to HUVEC by inhibiting VCAM-1 expression. These results demonstrate that GSPE is highly bioavailable and may serve as a potential therapeutic tool in protecting multiple target organs from structurally diverse drug- and chemical-induced toxicity.